AB0069 MEASUREMENT OF IL-33/ST2 AXIS IN PATIENTS WITH ACTIVE RHEUMATOID ARTHRITIS AND ITS ASSOCIATION WITH OTHER CYTOKINES IN SW982 AND U937 CELL LINES
Authors: M. Pai, V. Pai, P. Kumar, G. Badoni, S. Saha, P. Gupta
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> An inflammatory maelstrom is a characteristic feature of autoimmune diseases including Rheumatoid Arthritis (RA). IL-33, a member of IL-1 cytokine family has been studied in animal models with induced RA and a few studies in RA patients. These studies have shown the exacerbation of IL-33, however little is known about its pathogenesis and association with other cytokines in vivo and in vitro. <h3>Objectives</h3> This study evaluated the expression of IL33 and ST2 in serum and Peripheral blood mononuclear cells (PBMC's) of active RA patients, in addition to other cytokines (IL-13, IL-5, IL-17) and biochemical parameters. TNF-alpha stimulated and synovial fluid (SF) conditioned growth medias were used to grow cell lines (SW982, U937) so as to have ambient physio-pathological microenvironment during experiments, typically of RA and then the levels of IL-33 were evaluated. Association of IL-33 with expression of LTB4 was also studied. <h3>Methods</h3> PBMC's and sera from 50 active RA patients and 50 healthy individuals were obtained. IL-33 and ST2 were measured using Real Time PCR with specific primers including internal controls. Sandwich ELISA was done for studying protein expressions. Cell lines were grown RPMI-1649 + 10%FBS for U937 and DMEM + Glutamine + 10% FBS for SW982 cells at 5% CO<sub>2</sub>. <h3>Results</h3> IL-33 and ST2 mRNA expression was significantly higher in patients with active RA, with estimated fold change of 3.533(ΔΔCt = -1.82121) and 4.172 (ΔΔCt = -2.15229) respectively, as compared to controls. ELISA results revealed mean value of IL-33 to be significantly higher in RA patients than in controls (119.32±63.49 and 22.835±12.24 respectively). An increase in serum protein of ST2 was also observed in RA patients as compared to healthy controls (2434.328±834.60 and 705.22±310.05). Out of the total (n<sup>RA</sup> =50), 35 RA patients showed a hike in serum and mRNA expression of IL-33 which also showed correlation with other disease biomarkers like anti-CCP, CRP, DAS-28 CRP and HAQ-DI. Significant increase in serum protein of IL-17 (1542.112±120.21) and IL-5 (624.21±78.09) were observed. SF stimulated SW982 cells showed a 1.8 fold increase in levels of IL-33 as compared to cells which were stimulated with TNF alpha alone. Further monocytic cell line was treated with recombinantanti human IL-33 antibody and showed direct association with levels of Leukotriene B4 (LTB4) expression. <h3>Conclusion</h3> Elevated IL-33 and ST2 in both serum and PBMC's of RA patients showed correlation with the disease activity. Therefore, IL-33/ST2L signalling represents to be involved in the pathogenesis of RA and also indicates it to be a promising biomarker. Synovial fluid gives a natural microenvironment to cells in in-vitro studies showing increased expression of cytokines. This is the first study that shows a direct relation with IL-33 and LTB4 expression, thus proving the chemoattractive role of LTB4. <h3>References</h3> [1]Palmer G, Talabot-Ayer D, Lamacchia C, et al. Inhibition of interleukin-33 signaling attenuates the severity of experimental arthritis. <i>Arthritis Rheum.</i> 2009;60(3):738-749. doi:10.1002/art.24305 [2]Verri WA Jr, Souto FO, Vieira SM, et al. IL-33 induces neutrophil migration in rheumatoid arthritis and is a target of anti-TNF therapy. <i>Ann Rheum Dis.</i> 2010;69(9):1697-1703. [3]Duan L, Chen J, Gong F, Shi G. The role of IL-33 in rheumatic diseases. <i>Clin Dev Immunol</i>. 2013;2013:924363. [4]Miyabe Y etal. LTb4 and BLT1 in inflammatory arthritis. <i>Seminars in Immunology</i>. 2017; 33:52-57. <h3>Acknowledgements</h3> I deeply acknowledge Department of Science and Technology (DST) to award me with WoS A fellowship and grant to carry on my present research. <h3>Disclosure of Interests</h3> None Declared.